Abstract
Background
Human papillomaviruses (HPVs) remain a serious world health problem due to their association with anogenital/oral cancers and warts. While over 100 HPV types have been identified, a subset is associated with malignancy. HPV16 and 18 are the most prevalent oncogenic types, while HPV6 and 11 are most commonly responsible for anogenital warts. While other quantitative PCR (qPCR) assays detect oncogenic HPV, there is no single tube assay distinguishing the most frequent oncogenic types and the most common types found in warts.
Results
A Sybr Green-based qPCR assay was developed utilizing degenerate primers to the highly conserved HPV E1 theoretically detecting any HPV type. A single tube multiplex qPCR assay was also developed using type-specific primer pairs and TaqMan probes that allowed for detection and quantitation of HPV6,11,16,18. Each HPV type was detected over a range from 2 &#215; 101 to 2 &#215; 106copies/reaction providing a reliable method of quantitating type-specific HPV in 140 anogenital/cutaneous/oral benign and malignant specimens. 35 oncogenic and low risk alpha genus HPV types were detected. Concordance was detected in previously typed specimens. Comparisons to the gold standard detected an overall sensitivity of 89% (95% CI: 77% - 96%) and specificity of 90% (95%CI: 52% - 98%).
Conclusion
There was good agreement between the ability of the qPCR assays described here to identify HPV types in malignancies previously typed using standard methods. These novel qPCR assays will allow rapid detection and quantitation of HPVs to assess their role in viral pathogenesis.

Creator

Seaman, William TAffiliation: Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill

Andrews, ElizabethAffiliation: Department of Dental Ecology, School of Dentistry, University of North Carolina at Chapel Hill
College of Dental Medicine, Western University of Health Sciences, Pomona, CA, USA

Couch, MarionAffiliation: Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill

Palefsky, JoelAffiliation: Department of Medicine, University of California, San Francisco, San Francisco, CA, USA

Deal, Allison MAffiliation: Biostatistics Core Facility, University of North Carolina at Chapel Hill

Webster-Cyriaque, JenniferAffiliation: Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill
Department of Dental Ecology, School of Dentistry, University of North Carolina at Chapel Hill
Department of Microbiology and Immunology, University of North Carolina at Chapel Hill

Title

Detection and quantitation of HPV in genital and oral tissues and fluids by real time PCR